More than 20 ingredients of PDM-756, a partly defined, serum-free medium for axenic cultivation of E. histolytica are essential for growth for the ameba. In the presence of cysteine and NH4Cl no other amino acids appear to be required for growth. Radiolabeling studies show amebae grown in PDM-756 are metabolically similar to those grown in standard crude media implying their usefulness for metabolic studies of the parasite. Southern hybridization and electrophoretic migration of ribosomal DNA through ethidium bromide provided evidence that rRNA genes of E. histolytica are located on extrachromosomal circles. Direct evidence of circular DNA molecules was obtained by electron microscopy. Additionally, two-dimensional agarose gel electrophoretic studies shown the presence of replicating intermediates of the ribosomal DNA circles. Antigens recognized by humoral responses in patients infected with E. histolytica were identified and characterized by metabolic labeling followed by immunoprecipitation. A major antigen (31-36 kd) was detected with all patients' sera, including sera from cyst passers. Several monoclonal antibodies produced against cell surface and soluble antigens of E. histolytica recognized antigens from all tested isolates of E. histolytica, but not from other species of Entamoeba, including the non-pathogenic E. histolytica-like Laredo type ameba from humans.